The goal of this project is to elucidate some features of the structure of the Escherichia coli ribosome. In addition the mechanism of tRNA and mRNA interaction with ribosomes will be studied. Information is also sought on the mode of action of several antibiotics on bacterial ribosomes. These include erythromycin, streptomycin and chloramphenicol. If successful, the experiments planned will clearly establish the location of ribosome bound tRNA's and will uncover the details of conformational changes which accompany protein synthesis. Most work will involve the use of specifically attached fluorescent probes. The major emphasis is singlet-singlet energy transfer measurements of distances between specific sites. Polarization and quenching studies will also be performed to examine site flexibility and accessibility. Fluorescence will be used to monitor kinetics and equilibria involved in various discrete steps in protein synthesis. Electron microscopy using antibodies elicited against fluorescent dyes will be used to locate their position on the ribosome. Affinity labeling studies will identify components of functional sites. Lactoperoxidase iodination will examine gross changes in ribosome morphology introduced by the binding of subunits, factors, ligands, or antibodies. While restricted to bacterial ribosomes these studies should help to identify efficient strategies for eventual definitive study of the even more complicated mammalian ribosome.